Heterocyclic Substituted Acardite Derivate and Application Thereof

ABSTRACT

This present invention discloses a heterocyclic substituted acardite derivate and application thereof, namely compounds in the general formula (1) or the general formula (2) or pharmaceutically acceptable salts thereof, wherein A is monosubstituted or polysubstituted quinoline, isoquinoline, quinazoline, pyrrole or pyrimidine, and the substituent is halogen, C 1-5 alkyl, C 1-5 haloalkyl, C 1-5 alkoxy, C 1-5 haloalkoxy, C 1-5 alkylamino, C 1-5 haloalkylamino, amino or nitryl; R 1  is C 1-5 alkyl; R 2  is one or more selected from hydrogen, halogen, alkyl, alkoxy, haloalkyl or haloalkoxy; and R 3  is one or more selected from hydrogen, halogen, alkyl, alkoxy, haloalkyl or haloalkoxy. The compound of the present invention and the pharmaceutically acceptable salt thereof can be used for treating tumor or leukemia.

FIELD OF THE INVENTION

This present invention relates to an aromatic heterocyclic substitutedacardite derivate and application thereof In addition, the presentinvention relates to application of aromatic heterocyclic substitutedacardite derivate and pharmaceutically acceptable salts thereof in thetreatment of tumor or leukemia.

BACKGROUND OF THE INVENTION

With better understanding of the tumor molecular mechanisms, theresearch on the targeted therapy of the tumor moleculars has achievedimportant advance. Protein kinase inhibitor is one of newly developedtargeted therapy drugs, which affects the survival, proliferation anddisease progression of tumor cells through blocking the intra-cellularmolecular transduction pathway. Raf kinases play a crucial role in thesignal transduction pathway of Raf/MEK/ERK. Although the function of theRaf kinase in normal tissues is not yet understood, but the existingbasic and clinical research results have shown that the upregulation ofRaf gene and overexpression of its protein are present in various solidtumors, including renal cell carcinoma, hepatocellular carcinoma,melanoma and non-small cell lung cancer. Currently, more and more singletarget point and multi-target point therapy drugs for Raf kinases aresuccessfully developed and applied clinically, for example, sorafeniband erlotinib have achieved good clinical results, and the anti-tumortherapy has came into the “molecular targeted therapy” era.CN200810129360.6 disclosed that a kind of aromatic heterocyclicsubstituted acardite derivates with no substituent or only carbamyl inthe A ring have prospect of inhibiting specific tumors, and thepreliminary pharmacological experiments found that the effects of somecompouns are better than sorafenib.

SUMMARY OF THE INVENTION

The objective of the present invention is to provide an aromaticheterocyclic substituted acardite derivate having more medicinal valuethrough structural modification based on the existing technology. Afterthe present invention adds specific substituents in the A ring,especially adding substituents in the quinazoline, pyrrole or pyrimidinerings, the inhibitory activity and selectivity of the compounds tospecific tumors are greatly increased, and the absorptivity andutilization rate of the compounds are increased and the toxic sideeffects are reduced. The objective of the present invention is furtherto provide application of the compound or pharmaceutically acceptablesalts thereof in the treatment of tumor or leukemia. The heterocyclicsubstituted acardite derivate of the present invention can berepresented by the following formulas [1] and [2]:

-   -   wherein,

A is monosubstituted or polysubstituted quinoline, isoquinoline,quinazoline, pyrrole or pyrimidine, preferably monosubstituted orpolysubstituted quinazoline, pyrrole or pyrimidine, further preferablymonosubstituted or polysubstituted quinazoline; the substituent ishalogen, alkyl, haloalkyl, alkoxy, haloalkoxy, alkylamino,haloalkylamino, amino or nitryl, preferably halogen, C₁₋₅ alkyl, C₁₋₅haloalkyl, C₁₋₅ alkoxy, C₁₋₅ haloalkoxy, C₁₋₅ alkylamino,C₁₋₅haloalkylamino, amino or nitryl, more preferably halogen, C₁₋₅alkyl,C₁₋₅haloalkyl, C₁₋₅alkoxy, C₁₋₅haloalkoxy, amino or nitryl; still morepreferably halogen, amino, C₁₋₅alkyl or C₁₋₃alkoxy, particularlypreferably Cl, Br, F, amino, methoxy, methyl, ethyl, propyl, isopropyl,butyl or t-butyl in the present invention.

R₁ is alkyl, more preferably C₁₋₅alkyl, most preferably methyl, ethyl,propyl and isopropyl.

R₂ is one or more selected from hydrogen, halogen, alkyl, alkoxy,haloalkyl or haloalkoxy; preferably one or more selected from hydrogen,halogen, C₁₋₅alkyl, C₁₋₅alkoxy, C₁₋₅haloalkyl or C₁₋₅haloalkoxy, mostpreferably one or more selected from H, Cl, Br, F, methoxy, ethoxy,propoxy, methyl, ethyl, propyl, isopropyl, butyl, t-butyl ortrifluoromethyl.

R₃ is one or more selected from hydrogen, halogen, alkyl, alkoxy, C₁₋₅haloalkyl or C₁₋₅haloalkoxy, preferably one or more selected fromhydrogen, halogen, C₁₋₅alkyl, C₁₋₅alkoxy, C₁₋₅haloalkyl orC₁₋₅haloalkoxy, most preferably one or more selected from H, Cl, Br, F,methoxy, ethoxy, propoxy, methyl, ethyl, propyl, isopropyl, butyl,t-butyl or trifluoromethyl.

The pharmaceutically acceptable salts of the compound in the presentinvention are selected from:

-   a) basic salts of inorganic acids and organic acids, the described    acid is selected from hydrochloric acid, hydrobromic acid, sulfuric    acid, phosphoric acid, mesylate, trifluoromethanesulfonic acid,    benzene sulfonic acid, paratoluenesulfonic acid, 1-naphthalene    sulfonic acid, 2-naphthalene sulfonic acid, acetic acid,    trifluoroacetic acid, malic acid, tartaric acid, citric acid, lactic    acid, oxalic acid, succinic acid, fumaric acid, maleic acid, benzoic    acid, salicylic acid, phenylacetic acid or almonds acid;-   b) acid salts of organic and inorganic base, the described cation is    selected from alkali metal cation, alkaline earth metal cation,    ammonium cation, aliphatic-substituted ammonium cation or    aromatic-substituted ammonium cation.

Preparation of the Compound of Formula 1

Method 1: the target compound is obtained from substituted heterocyclic2-carboxylate as starting materials through acyl chlorination,aminoalkylation, two-step condensation and salt forming reaction and theroute is as follows:

Method 2: the target compound is obtained from substituted heterocyclic2-carboxylate as starting materials through acyl chlorination,aminoalkylation, condensation and salt forming reaction and the route isas follows:

Preparation of the Compound of Formula 2

The target compound is obtained from halogen substituted heterocyclic asstarting materials through two-step condensation and salt formingreaction and the route is as follows:

The substituents A, R₁, R₂ and R₃ in the above menthioned reactionroutes have the above described meanings.

The beneficial effects of the present invention are as follows:

The derivatives of the present invention have raf kinase inhibitoryactivity. The action mechanism of this compound is that this compoundaffects the survival, proliferation and disease progression of tumorcells through inhibiting raf kinase and blocking the ras protein signaltransduction connection, thereby inhibiting the growth of achiblastomas,such as malignant tumors (for example, bladder cancer, lung cancer,pancreatic cancer), myelopathy (for example, myelogenous leukemia) oradenoma (for example, villous adenoma of colon).

The experiment results have shown that the compound with specialsubstituents added in A ring in the present invention has strongerantitumor activity compared with the previously disclosed compound withno substituent or only carbamyl in A ring, which is obviously strongerthan Sorafenib in the effects of tumor cell metastasis and tumorangiogenesis. The test on normal human umbilical vein endothelial cellsfound that this part of the compounds have lower toxicity to normalhuman cells, such as endothelial cell, which are safe and reliable, butwhich can inhibit the tumor angiogenesis to achieve anti-tumor activity.In vivo nude mice transplanted model experiment proved that the compoundof the present invention has inhibitory effects to human liver andkidney cancer and the effects are stronger than Sorafenib, which hasmore obvious effects on lung cancer and the effects are significantlybetter than the positive control drug Sorafenib. The results show thatthe compound of the present invention or pharmaceutically acceptablesalts thereof can be used in the drugs for the treatment of cancer orleukemia, particularly drugs used for treating lung cancer.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

The melting point was measured by the electric melting point instrumentand the thermometer was not corrected; the elemental analyzer wasFoss-Heraeus type; and the mass spectrograph was electrospray ionizationmass spectrometry.

A: preparing the aromatic heterocyclic substituted acardite derivatehaving the general formula 1 accroding to method 1

Embodiment 1 Preparation of 4-Chloro-6-Methoxyquinolinyl-2-CarbonylChloride

50g of 4-hydroxy-6-methoxy-2-quinolinecarboxylic acid and 100 m1 ofthionyl chloride were added into a three-necked flask, heated andrefluxed for 17 hours until the reaction finished. The filtrate wasadded with toluene and concentrated under vacuum to obtain yellow solid,namely 4-chloro-6-methoxyquinolinyl-2-carbonyl chloride, with dry weightof 50g.

Embodiment 2 Preparation of 4-Chloro-7-Fluoroquinazolinyl-2-CarbonylChloride

Prepared from 4-hydroxy-7-fluoro-2-quinazolinecarboxylic acid withreference to the method of embodiment 1.

Embodiment 3 Preparation of 4-Methoxy-5-Chloropyrimidine-2-CarbonylChloride

Prepared from 4-methoxy-5-hydroxy-2-pyrimidinecarboxylic acid withreference to the method of embodiment 1.

Embodiment 4 Preparation of 4-Chloro-7-Amino-Isoquinolyl-2-CarbonylChloride

Prepared from 4-hydroxy-7-amino-2-quinazolinecarboxylic acid withreference to the method of embodiment 1.

Embodiment 5 Preparation of 5-Methyl-4-Chloropyrrolyl-2-CarbonylChloride

Prepared from 5-methyl-4-hydroxy-2-pyrrolecarboxylic acid with referenceto the method of embodiment 1.

Embodiment 6 Preparation of 4-Chloro-6-Methoxyl-N-methyl-2-QuinolinylFormamide

10 g of 4-chloro-6-methoxyquinolinyl-2-carbonyl chloride (obtained fromembodiment 1) was reacted with 200 ml of 2M methylamine ethanol solutionunder 0° C. for 36 hours until the reaction finished. The solvent wasevaporated under vacuum and the residues were added with water followedby stirring evenly. Ethyl acetate was added for extracting and the ethylacetate layer was dried with anhydrous sodium sulfate. The ethyl acetatelayer was removed under vacuum to obtain 9 g of4-chloro-6-methoxy-N-methyl-2-quinoline carboxamide.

Embodiment 7 Preparation of 4-Chloro-7-Fluoro-N-Methyl-2-QuinazolineMethanamide

Prepared from 4-chloro-7-fluoroquinazolinyl-2-carbonyl chloride withreference to the method of embodiment 6.

Embodiment 8 Preparation of4-Methoxyl-5-Chloro-N-Methyl-2-Pyrimidinecarboxamide

Prepared from 4-methoxyl-5-chloropyrimidinyl-2-carbonyl chloride withreference to the method of Embodiment 6.

Embodiment 9 Preparation of4-Chloro-7-Amino-N-Methyl-2-Isoquinolinecarboxamide

Prepared from 4-chloro-7-aminoisoquinolyl-2-carbonyl chloride withreference to the method of embodiment 6.

Embodiment 10 Preparation of 5-Methyl-4-Chloro-N-Methyl-2-PyrroleCarboxamide

Prepared from 5-methyl-4-chloropyrryl-2-carbonyl chloride with referenceto the method of embodiment 6.

Embodiment 11 Preparation of

4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl) oxy))aniline 10 gof 4-chloro-6-methoxyl-N-methyl-2-quinoline carboxamide (obtained fromembodiment 6) was dissolved in DMF, added with 20 g of potassiumtert-butylate and 10 g of 4-aminophenol and kept at 70° C. under theprotection of nitrogen for 8 hours. After the reaction finished, thereaction solution was poured into 250 ml of ethyl acetate and 250 ml ofsaturated saline solution and stirred evenly for separation. The watersolution was extracted with ethyl acetate again. The ethyl acetate layerwas washed with saturated saline solution and dried with anhydroussodium sulfate. The solvent was evaporated and removed under vacuum toobtain 6 g of4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))aniline.

Embodiment 12 Preparation of4-(2-(N-Methylaminoformoxyl)-4-(7Fluoroquinazolinyl) Oxy)Aniline

Prepared from 4-chloro-7-fluoro-N-methyl-2-quinazoline methanamide withreference to the method of embodiment 11.

Embodiment 13 Preparation of

4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl) oxy)anilinePrepared from 4-methoxyl-5-chloro-N-methyl-2-pyrimidinecarboxamide withreference to the method of embodiment 11.

Embodiment 14 Preparation of4-(2-(N-Methylaminoformoxyl)-4-(7-Amino-Isoquinolyl) Oxy)Aniline

Prepared from 4-chloro-7-amino-N-methyl-2-isoquinolinecarboxamide withreference to the method of embodiment 11.

Embodiment 15 Preparation of4-(2-(N-Methylaminoformoxyl)-4-(5-Methyl-Pyrryl)Oxy) Aniline

Prepared from 5-methyl-4-chloro-N-methyl-2-pyrrole carboxamide withreference to the method of embodiment 11.

Embodiment 16 Synthesis of 4-Chloro-3-(Trifluoromethyl)Phenyl Isocyanate

20 g of 4-chloro-3-(trifluoromethyl)aniline was mixed with 100 mlbenzene, added with 20g of diphosgene and refluxed for 12 hours. Thereaction solution was added with toluene, and the solvent was evaporatedand removed under vacuum to obtain the product4-chloro-3-(trifluoromethyl)phenyl isocyanate.

Embodiment 17 Synthesis of 4-Bromo-3-(Trifluoromethyl)Phenyl Isocyanate

Prepared from 4-bromo-3-(trifluoromethyl)aniline with reference to themethod of embodiment 16.

Embodiment 18 Synthesis of 4-Fluoro-3-(Trifluoromethyl)Phenyl Isocyanate

Prepared from 4-fluoro-3-(trifluoromethyl)aniline with reference to themethod of embodiment 16.

Embodiment 19 Synthesis of Compound 1

7 g of 4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))aniline(obtained from embodiment 11), 5 g of 4-chloro-3-(trifluoromethyl)phenylisocyanate (obtained from embodiment 16) and 50 ml of methylenedichloride were stirred at room temperature for 24 hours, and thecrystals were separated out followed by air pump filtration andcollection to obtain

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))phenyl)urea.

Embodiment 20 Synthesis of Compound 2

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl) oxy))anilineanilineand

4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the method ofEmbodiment 19.

Embodiment 21 Synthesis of Compound 3

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))aniline and

4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the method ofEmbodiment 19.

Embodiment 22 Synthesis of Compound 4

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazolinyl)oxy) phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazolinyl)oxy)aniline(obtained from embodiment 12) and 4-chloro-3-(trifluoromethyl)phenylisocyanate according to the method of Embodiment 19.

Embodiment 23 Synthesis of Compound 5

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazolinyl)oxy) phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazolinyl)oxy)aniline and

4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the method ofEmbodiment 19.

Embodiment 24 Synthesis of Compound 6

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazolinyl)oxy) phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazolinyl)oxy)aniline and

4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the method ofEmbodiment 19.

Embodiment 25 Synthesis of Compound 7

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl)oxy) phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl)oxy)aniline(obtained from embodiment 13) and 4-chloro-3-(trifluoromethyl)phenylisocyanate according to the method of Embodiment 19.

Embodiment 26 Synthesis of Compound 8

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl)oxy) phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl)oxy)aniline and

4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the method ofEmbodiment 19.

Embodiment 27 Synthesis of Compound 9

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl)oxy) phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl)oxy)aniline and

4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the method ofEmbodiment 19.

Embodiment 28 Synthesis of Compound 10

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoquinolyl)oxy)phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoquinolyl)oxy)aniline(obtained from embodiment 14) and 4-chloro-3-(trifluoromethyl)phenylisocyanate according to the method of Embodiment 19.

Embodiment 29 Synthesis of Compound 11

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoquinolyl)oxy)phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-isoquinolyl)oxy)aniline and

4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the method ofEmbodiment 19.

Embodiment 30 Synthesis of Compound 12

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoquinolyl)oxy)phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-isoquinolyl)oxy)aniline and

4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the method ofEmbodiment 19.

Embodiment 31 Synthesis of Compound 13

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(5-methyl-pyrryl)oxy)phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(5-methyl-pyrryl)oxy)aniline (obtainedfrom embodiment 15) and 4-chloro-3-(trifluoromethyl)phenyl isocyanateaccording to the method of Embodiment 19.

Embodiment 32 Synthesis of Compound 14

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(5-methyl-pyrryl)oxy)phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(5-methyl-pyrryl)oxy)aniline and

4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the method ofEmbodiment 19.

Embodiment 33 Synthesis of Compound 15

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(5-methyl-pyrryl)oxy)phenyl)urea was prepared from

4-(2-(N-methylaminoformoxyl)-4-(5-methyl-pyrryl)oxy)aniline and

4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the method ofEmbodiment 19.

Embodiment 34 Synthesis of

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))phenyl)urea mesylate

10 g of

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))phenyl)urea free base was dissolved in 300 ml of ether andadded with methanesulfonic acid/ethanol solution in drops at roomtemperature until pH=2, and white crystal was precipitated followed byair pump filtration and collection to obtain

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))phenyl)urea mesylate.

Embodiment 35 Synthesis of Pharmaceutically Acceptable Salts of

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))phenyl)urea

With reference to the method of embodiment 34, fluoromethanesulfonicacid/ethanol solution, benzene sulfonic acid/ethanol solution,p-toluenesulfonic acid/ethanol solution,

1-naphthalenesulfonic acid/ethanol solution, 2-naphthalenesulfonicacid/ethanol solution, acetic acid/ethanol solution, trifluoroaceticacid/ethanol solution, malic acid/ethanol solution, tartaricacid/ethanol solution, citric acid/ethanol solution, lactic acid/ethanolsolution, oxalic acid/ethanol solution, succinic acid/ethanol solution,fumaric acid/ethanol solution, maleic acid/ethanol solution, benzoicacid/ethanol solution, salicylic acid/ethanol solution, phenylaceticacid/ethanol solution or mandelic acid/ethanol solution were added indrops to synthesize trifluoromethylsulfonate, benzene sulfonate,tosilate, 1-naphthalenesulfenesulfonate, 2-naphthalenesulfenesulfonate,acetate, trifluoroactate, malate, tartrate, citrate, lactate, oxalate,succinate, fumarate, maleate, benzoate, salicylate, phenylacetate ormandelate of N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))phenyl)urea.

The pharmaceutically acceptable salts of compounds 2-15 can be alsosynthesized according to the above mentioned method.

B: preparing the aromatic heterocyclic substituted acardite derivatehaving the general formula 1 accroding to method 2

Embodiment 36 Synthesis of

N-(4-chloro-3 -(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea

20 g of 4-chloro-3-(trifluoromethyl)phenyl isocyanate (obtained fromembodiment 16), 15 g of 4-aminophenol and 500 ml of dichloromethane werestirred at room temperature for 2 h, and the crystal was precipitated,followed by air pump filtration and collection, washing withdichloromethane and vacuum drying to obtain

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea.

Embodiment 37 Synthesis of

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea

Prepared from 4-bromo-3-(trifluoromethyl)phenyl isocyanate (obtainedfrom embodiment 17) with reference to embodiment 36.

Embodiment 38 Synthesis of

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea

Prepared from 4-fluoro-3-(trifluoromethyl)phenyl isocyanate (obtainedfrom embodiment 18) with reference to embodiment 36.

Embodiment 39 Synthesis of

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))phenyl)urea (compound 1)

10 g of N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea(obtained from embodiment 36), 8.2 g of4-chloro-6-methoxyl-N-methyl-2-quinoline carboxamide (obtained fromembodiment 6) and 50 ml dichloromethane were stirred at room temperaturefor 24 h, and the crystal was precipitated, followed by air pumpfiltration and collection to obtain 12 g ofN-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))phenyl)urea.

Embodiment 40 Synthesis of Compound 2

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))phenyl)urea was prepared from

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-6-methoxyl-N-methyl-2-quinoline carboxamide according to themethod of Embodiment 39.

Embodiment 41 Synthesis of Compound 3

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(6-methoxyquinolinyl)oxy))phenyl)urea was prepared from

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-6-methoxyl-N-methyl-2-quinoline carboxamide according to themethod of Embodiment 39.

Embodiment 42 Synthesis of Compound 4

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazolinyl)oxy) phenyl)urea was prepared from

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-7-fluoro-N-methyl-2-quinazoline methanamide (obtained fromembodiment 7) according to the method of Embodiment 39.

Embodiment 43 Synthesis of Compound 5

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazolinyl)oxy)phenyl)urea was prepared from

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-7-fluoro-N-methyl-2-quinazoline methanamide according to themethod of Embodiment 39.

Embodiment 44 Synthesis of Compound 6

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7fluoroquinazolinyl)oxy)phenyl)urea was prepared from

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-7-fluoro-N-methyl-2-quinazoline methanamide according to themethod of Embodiment 39.

Embodiment 45 Synthesis of Compound 7

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl)oxy) phenyl)urea was prepared from

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-methoxyl-5-chloro-N-methyl-2-pyrimidinecarboxamide (obtained fromembodiment 8) according to the method of Embodiment 39.

Embodiment 46 Synthesis of Compound 8

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl)oxy)phenyl)urea was prepared from

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and4-methoxyl-5-chloro-N-methyl-2-pyrimidinecarboxamide according to themethod of Embodiment 39.

Embodiment 47 Synthesis of Compound 9

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-5-(4-methoxypyrimidinyl)oxy)phenyl)urea was prepared from

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-N-methyl-2-pyrimidinecarboxamide according to the method ofEmbodiment 39.

Embodiment 48 Synthesis of Compound 10

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoquinolyl)oxy)phenyl)urea was prepared from

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-7-amino-N-methyl-2-isoquinolinecarboxamide (obtained fromembodiment 9) according to the method of Embodiment 39.

Embodiment 49 Synthesis of Compound 11

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoquinolyl)oxy)phenyl)urea was prepared from

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-7-amino-N-methyl-2-isoquinolinecarboxamide according to themethod of Embodiment 39.

Embodiment 50 Synthesis of Compound 12

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(7-amino-isoquinolyl)oxy)phenyl)urea was prepared from

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

4-chloro-7-amino-N-methyl-2-isoquinolinecarboxamide according to themethod of Embodiment 39.

Embodiment 51 Synthesis of Compound 13

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(2-methyl-pyrryl)oxy)phenyl)urea was prepared from

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

2-methyl-4-chloro-N-methyl-2-pyrrole carboxamide (prepared fromembodiment 10) according to the method of Embodiment 39.

Embodiment 52 Synthesis of Compound 14

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(2-methyl-pyrryl)oxy)phenyl)urea was prepared from

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

2-methyl-4-chloro-N-methyl-2-pyrrole carboxamide according to the methodof Embodiment 39.

Embodiment 53 Synthesis of Compound 15

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylaminoformoxyl)-4-(2-methyl-pyrryl)oxy)phenyl)urea was prepared from

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-hydroxylphenyl)urea and

2-methyl-4-chloro-N-methyl-2-pyrrole carboxamide according to the methodof Embodiment 39.

C: preparing the aromatic heterocyclic substituted acardite derivatehaving the general formula 2 accroding to method 3

Embodiment 54 Preparation of 4-(4-(6-Methoxyquinolinyl)Oxy))Aniline

8 g of 4-chloro-6-methoxyquinoline was dissolved in DMF, added with 20 gof potassium tert-butylate and 10 g of 4-aminophenol and reacted underthe protection of nitrogen at 70° C. for 8 hours. After the end of thereaction, the reaction liquid was poured into 250 ml of ethyl acetateand 250 ml of saturated salt water and mixed evenly followed by liquidseparation. The water solution was extracted with ethyl acetate.

The ethyl acetate layer was added with saturated salt water for washingand dried by anhydrous sodium sulfate. The solvent was evaporated undervacuum to obtain 6 g of 4-(4-(6-methoxyquinolinyl)oxy))aniline.

Embodiment 55 Preparation of 4-(4-(7Fluoroquinazolinyl)Oxy)Aniline

Prepared from 4-chloroquinazoline with reference to the method ofembodiment 54.

Embodiment 56 Preparation of 4-(5-(4-Methoxypyrimidinyl)Oxy)Aniline

Prepared from 5-chloro-4-methoxypyrimidine with reference to the methodof embodiment 54.

Embodiment 57 Preparation of 4-(4-(7-Amino-Isoquinolyl)Oxy)Aniline

Prepared from 4-chloro-7-aminoisoquinoline with reference to the methodof embodiment 54.

Embodiment 58 Preparation of 4-(4-(2-Methyl-Pyrryl)Oxy)Aniline

Prepared from 4-chloro-2-methylpyrrol with reference to the method ofembodiment 54.

Embodiment: 59 Preparation of4-(4-(6-Methoxyl-7-Fluoro-Quinolinyl)Oxy)Aniline

Prepared from 4-chloro-6-methoxyl-7-fluoro-quinoline with reference tothe method of embodiment 54.

Embodiment: 60 Preparation of4-(4-(6-Methyl-7-Fluoro-Quinolinyl)Oxy))Aniline

Prepared from 4-chloro-6-methyl-7-fluoro-quinazoline with reference tothe method of embodiment 54.

Embodiment 61 Synthesis of 4-Chloro-3-(Trifluoromethyl)Phenyl Isocyanate

100 ml of diphosgene is mixed with 20 g of4-chloro-3-(trifluoromethyl)aniline and refluxed for 12 hours. Thereaction liquid was added into toluene, and the solvent was evaporatedunder vacuum to obtain the product 4-chloro-3-(trifluoromethyl)phenylisocyanate.

Embodiment 62 Synthesis of 4-Bromo-3-(Trifluoromethyl)Phenyl Isocyanate

Prepared from 4-bromo-3-(trifluoromethyl)aniline with reference to themethod of embodiment 61.

Embodiment 63 Synthesis of 4-Fluoro-3-(Trifluoromethyl)Phenyl Isocyanate

Prepared from 4-fluoro-3-(trifluoromethyl)aniline with reference to themethod of embodiment 61.

Embodiment 64 Synthesis of 4-Chloro-3-Ethylphenyl Isocyanate

Prepared from 4-chloro-3-ethylaniline with reference to the method ofembodiment 61.

Embodiment 65 Synthesis of 4-Ethyl-3-Trifluoromethyl

Prepared from 4-ethyl3-trifluoromethylaniline with reference to themethod of embodiment 61.

Embodiment 66 Synthesis of

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-quinolinyl)oxyphenyl)urea(compound 16) 7 g of4-(4-(6-methoxyquinolinyl)oxy))aniline4-(4-quinolinyl)oxyaniline(prepared from embodiment 54), 5 g of 4-chloro-3-(trifluoromethyl)phenylisocyanate (prepared from embodiment 61) and 50 ml of methylenedichloride were mixed and reacted at room temperature for 24 hours, andthe crystal was precipitated, followed by air pump filtration andcollection to obtain

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy))phenyl)urea.

Embodiment 67 Synthesis of Compound 17

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy))phenyl)ureawas prepared from 4-(4-(6-methoxyquinolinyl)oxy))aniline and4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the method ofEmbodiment 66.

Embodiment 68 Synthesis of Compound 18

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy))phenyl)ureawas prepared from 4-(4-(6-methoxyquinolinyl)oxy))aniline and4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the methd ofEmbodiment 66.

Embodiment 69 Synthesis of Compound 19

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7fluoroquinazolinyl)oxy)phenyl)ureawas prepared from 4-(4-(7fluoroquinazolinyl)oxy)aniline (prepared fromembodiment 55) and 4-chloro-3-(trifluoromethyl)phenyl isocyanateaccording to the methd of Embodiment 66.

Embodiment 70 Synthesis of Compound 20

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7fluoroquinazolinyl)oxy)phenyl)ureawas prepared from 4-(4-(7fluoroquinazolinyl)oxy)aniline and4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the methd ofEmbodiment 66.

Embodiment 71 Synthesis of Compound 21

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7fluoroquinazolinyl)oxy)phenyl)ureawas prepared from 4-(4-(7fluoroquinazolinyl)oxy)aniline and4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the methd ofEmbodiment 66.

Embodiment 72 Synthesis of Compound 22

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(5-(4-methoxypyrimidinyl)oxy)phenyl)ureawas prepared from 4-(5-(4-methoxypyrimidinyl)oxy)aniline (prepared fromembodiment 56) and 4-chloro-3-(trifluoromethyl)phenyl isocyanateaccording to the methd of Embodiment 66.

Embodiment 73 Synthesis of Compound 23

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(5-(4-methoxypyrimidinyl)oxy)phenyl)ureawas prepared from 4-(5-(4-methoxypyrimidinyl)oxy)aniline and4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the methd ofEmbodiment 66.

Embodiment 74 Synthesis of Compound 24

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(5-(4-methoxypyrimidinyl)oxy)phenyl)ureawas prepared from 4-(5-(4-methoxypyrimidinyl)oxy)aniline and4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the methd ofEmbodiment 66.

Embodiment 75 Synthesis of Compound 25

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)ureawas prepared from 4-(4-(7-amino-isoquinolyl)oxy)aniline (prepared fromembodiment 57) and 4-chloro-3-(trifluoromethyl)phenyl isocyanateaccording to the methd of Embodiment 66.

Embodiment 76 Synthesis of Compound 26

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)ureawas prepared from 4-(4-(7-amino-isoquinolyl)oxy)aniline and4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the methd ofEmbodiment 66.

Embodiment 77 Synthesis of Compound 27

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)ureawas prepared from 4-(4-(7-amino-isoquinolyl)oxy)aniline and4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the methd ofEmbodiment 66.

Embodiment 78 Synthesis of Compound 28

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(2-methyl-pyrryl)oxy)phenyl)ureawas prepared from 4-(4-(2-methyl-pyrryl)oxy)aniline (prepared fromembodiment 58) and 4-chloro-3-(trifluoromethyl)phenyl isocyanateaccording to the methd of Embodiment 66.

Embodiment 79 Synthesis of Compound 29

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(2-methyl-pyrryl)oxy)phenyl)ureawas prepared from 4-(4-(5-methyl-pyrryl)oxy)aniline and4-fluoro-3-(trifluoromethyl)phenyl isocyanate according to the methd ofEmbodiment 66.

Embodiment 80 Synthesis of Compound 30

N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(2-methyl-pyrryl)oxy)phenyl)ureawas prepared from 4-(4-(5-methyl-pyrryl)oxy)aniline and4-bromo-3-(trifluoromethyl)phenyl isocyanate according to the methd ofEmbodiment 66.

Embodiment 81 Synthesis of Compound 31

N-(4-chloro-3-ethylphenyl)-N′-(4-(4-(6-methoxyl-7-fluoro-quinolinyl)oxy)phenyl)ureawas prepared from 4-(4-(6-methoxyl-7-fluoro-quinolinyl)oxy)aniline(prepared from embodiment 59) and 4-chloro-3-ethylphenyl isocyanate(prepared from embodiment 64) according to the methd of Embodiment 66.

Embodiment 82 Synthesis of Compound 32

N-(4-ethyl-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyl-7-fluoro-quinolinyl)oxy)phenyl)ureawas prepared from 4-(4-(6-methoxyl-7-fluoro-quinolinyl)oxy)aniline(prepared from embodiment 59) and 4-ethyl-3-trifluoromethyl isocyanate(prepared from embodiment 65) according to the methd of Embodiment 66.

Embodiment 83 Synthesis of Compound 33

N-(4-chloro-3-ethylphenyl)-N′-(4-(4-(6-methyl-7-fluoro-quinolinyl)oxy))phenyl)ureawas prepared from 4-(4-(6-methyl-7-fluoro-quinolinyl)oxy))aniline(prepared from embodiment 60) and 4-chloro-3-ethylphenyl isocyanate(prepared from embodiment 64) according to the methd of Embodiment 66.

Embodiment 84 Synthesis of Compound 34

N-(4-ethyl-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methyl-7-fluoro-quinolinyl)oxy))phenyl)ureawas prepared from 4-(4-(6-methyl-7-fluoro-quinolinyl)oxy))aniline(prepared from embodiment 60) and 4-ethyl-3-trifluoromethyl isocyanate(prepared from embodiment 65) according to the methd of Embodiment 66.

Embodiment 85 Preparation of 4-Chloro-7-Nitrylquinoline-2-CarbonylChloride

Prepared from 4-hydroxyl-7-nitryl-2-quinoline carboxylic acid withreference to the method of embodiment 1.

Embodiment 86 Preparation of4-Chloro-7-Trifluoromethylquinazoline-2-Carbonyl Chloride

Prepared from 4-hydroxyl-7-trifluoromethyl-2-quinazoline carboxylic acidwith reference to the method of embodiment 1.

Embodiment 87 Preparation of 4-Chloro-7-Nitryl-N-Ethyl-2-QuinolineCarboxamide

Prepared from 4-chloro-7-nitrylquinoline-2-carbonyl chloride (preparedfrom embodiment 58) and 2M ethylamine ethanol solution with reference tothe method of embodiment 6.

Embodiment 88 Preparation of4-Chloro-7-Trifluoromethyl-N-Propyl-2-Quinazoline Methanamide

Prepared from 4-chloro-7-trifluoromethylquinazoline-2-carbonyl chloride(prepared from embodiment 86) and 2M propylamine ethanol solution withreference to the method of embodiment 6.

Embodiment 89 Preparation of

2-methyl-4-(2-(N-ethylcarbamyl)-4-(7-nitrylquinolinyl)oxy))aniline

Prepared from 4-chloro-7-nitryl-N-ethyl-2-quinoline carboxamide(prepared from embodiment 87) and 3-methyl-4-aminophenol with referenceto the method of embodiment 11.

Embodiment 90 Preparation of2-Methoxyl-4-(2-(N-Ethylcarbamyl)-4-(7-Nitrylquinolinyl)Oxy))Aniline

Prepared from 4-chloro-7-nitryl-N-ethyl-2-quinoline carboxamide(prepared from embodiment 87) and 3-methoxyl-4-aminophenol withreference to the method of embodiment 11.

Embodiment 91 Preparation of2-Fluoro-4-(2-(N-Propylcarbamyl)-4-(7-Trifluoromethylquinolinyl)Oxy))Aniline

Prepared from 4-chloro-7-trifluoromethyl-N-propyl-2-quinazolinemethanamide (prepared from embodiment 88) and 3-fluoro-4-aminophenolwith reference to the method of embodiment 11.

Embodiment 92 Preparation of2-Trifluoromethyl-4-(2-(N-Propylcarbamyl)-4-(7-Trifluoromethylquinolinyl)Oxy))Aniline

Prepared from 4-chloro-7-trifluoromethyl-N-propyl-2-quinazolinemethanamide (prepared from embodiment 88) and 4-amino-3-trifluoromethylphenol with reference to the method of embodiment 11.

Embodiment 93 Synthesis of 4-Chloro-3-Methoxyphenyl Isocyanate

Prepared from 4-chloro-3-methoxyaniline with reference to the method ofembodiment 61.

Embodiment 94 Preparation of Compound 35

N-(4-chloro-3-methoxyphenyl)-N′-(2-methyl-4-(2-(N-ethylcarbamyl)-4-(7-nitryl-quinolinyl)oxy)phenyl)urea was sythesized from2-methyl-4-(2-(N-ethylcarbamyl)-4-(7-nitrylquinolinyl)oxy))aniline(prepared from embodiment 89) and 4-chloro-3-methoxyphenyl isocyanate(prepared from embodiment 93) according to the method of Embodiment 19.

Embodiment 95 Preparation of Compound 36

N-(4-chloro-3-methoxyphenyl)-N′-(2-methoxyl-4-(2-(N-ethylcarbamyl)-4-(7-nitryl-quinolinyl)oxy)phenyl)urea was sythesized from2-methoxyl-4-(2-(N-ethylcarbamyl)-4-(7-nitrylquinolinyl)oxy))aniline(prepared from embodiment 90) and 4-chloro-3-methoxyphenyl isocyanate(prepared from embodiment 93) according to the method of Embodiment 19.

Embodiment 96 Preparation of Compound 37

N-(4-chloro-3-methoxyphenyl)-N′-(2-fluoro-4-(2-(N-propylcarbamyl)-4-(7-trifluoromethylquinolinyl)oxy))phenyl)urea was sythesized from2-fluoro-4-(2-(N-propylcarbamyl)-4-(7-trifluoromethylquinolinyl)oxy))aniline(prepared from embodiment 91) and 4-chloro-3-methoxyphenyl isocyanate(prepared from embodiment 93) according to the method of Embodiment 19.

Embodiment 97 Preparation of Compound 38

N-(4-chloro-3-methoxyphenyl)-N′-(2-trifluoromethyl-4-(2-(N-propylcarbamyl)-4-(7-trifluoromethylquinolinyl)oxy))phenyl)urea was sythesized from

2-trifluoromethyl-4-(2-(N-propylcarbamyl)-4-(7-trifluoromethylquinolinyl)oxy))aniline(prepared from embodiment 92) and 4-chloro-3-methoxyphenyl isocyanate(prepared from embodiment 93) according to the method of Embodiment 19.

Embodiment 98 Synthesis ofN-(4-Chloro-3-(Trifluoromethyl)Phenyl)-N′-(4-(4-(6-Methoxyquinolinyl)Oxy))Phenyl)UreaMesylate

10 g ofN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy))phenyl)ureafree base was dissolved in 300 ml of ether and added withmethanesulfonic acid/ethanol solution in drops at room temperature untilpH=2, and white crystal was precipitated followed by air pump filtrationand collection to obtainN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy))phenyl)ureamesylate.

Embodiment 99

Synthesis of Pharmaceutically Acceptable Salts ofN-(4-Chloro-3-(Trifluoromethyl)Phenyl)-N′-(4-(4-(6-Methoxyquinolinyl)Oxy))Phenyl)Urea

With reference to the method of embodiment 85, fluoromethanesulfonicacid/ethanol solution, benzene sulfonic acid/ethanol solution,p-toluenesulfonic acid/ethanol solution, 1-naphthalenesulfonicacid/ethanol solution, 2-naphthalenesulfonic acid/ethanol solution,acetic acid/ethanol solution, trifluoroacetic acid/ethanol solution,malic acid/ethanol solution, tartaric acid/ethanol solution, citricacid/ethanol solution, lactic acid/ethanol solution, oxalic acid/ethanolsolution, succinic acid/ethanol solution, fumaric acid/ethanol solution,maleic acid/ethanol solution, benzoic acid/ethanol solution, salicylicacid/ethanol solution, phenylacetic acid/ethanol solution or mandelicacid/ethanol solution were added in drops to synthesizetrifluoromethylsulfonate, benzene sulfonate, tosilate,1-naphthalenesulfenesulfonate, 2-naphthalenesulfenesulfonate, acetate,trifluoroactate, malate, tartrate, citrate, lactate, oxalate, succinate,fumarate, maleate, benzoate, salicylate, phenylacetate or mandelate ofN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-quinolinyl)oxyphenyl)urea.

The pharmaceutically acceptable salts of compounds 17-38 can be alsosynthesized according to the above mentioned method.

The compounds in table 1 to 14 were prepared according to methods of theabove mentioned embodiments, and the characteristics are shown in thefollowing tables.

TABLE 1 substituted quinoline derivatives

Compound Elementary Mass spectrum No. R₃ analysis Name M + 1 14-chloro-3- C: 57.3 N-(4-chloro-3-(trifluoromethyl)phenyl)-N′- 545.5trifluoromethyl H: 3.8 (4-(2-(N-methylaminoformoxyl)-4-(6- N: 10.3methoxyquinolinyl)oxy))phenyl)urea 2 4-fluoro-3- C: 59.2N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′- 529 trifluoromethyl H: 3.9(4-(2-(N-methylaminoformoxyl)-4-(6- N: 10.5methoxyquinolinyl)oxy))phenyl)urea 3 4-bromo-3- C: 53.0N-(4-bromo-3-(trifluoromethyl)phenyl)-N′- 590 trifluoromethyl H: 3.31(4-(2-(N-methylaminoformoxyl)-4-(6- N: 9.38methoxyquinolinyl)oxy))phenyl)urea

TABLE 2 substituted quinazoline derivatives

Compound Elementary Mass spectrum No. R₃ analysis Name M + 1 44-chloro-3- C: 54.1 N-(4-chloro-3-(trifluoromethyl)phenyl)-N′- 534.5trifluoromethyl H: 3.11 (4-(2-(N-methylaminoformoxyl)-4- N: 13.0 (7fluoroquinazolinyl)oxy)phenyl)urea 5 4-fluoro-3- C: 55.5N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′- 518 trifluoromethyl H: 3.30(4-(2-(N-methylaminoformoxyl)-4- N: 13.4 (7fluoroquinazolinyl)oxy)phenyl)urea 6 4-bromo-3- C: 49.7N-(4-bromo-3-(trifluoromethyl)phenyl)-N′- 579 trifluoromethyl H: 2.91(4-(2-(N-methylaminoformoxyl)-4- N: 12.2 (7fluoroquinazolinyl)oxy)phenyl)urea

TABLE 3 substituted pyrimidine derivatives

Compound Elementary Mass No. R₃ analysis Name spectrum m/e 7 4-chloro-3-C: 50.9 N-(4-chloro-3-(trifluoromethyl)phenyl)-N′- 496.5 trifluoromethylH: 3.48 (4-(2-(N-methylaminoformoxyl)-5-(4- N: 14.0methoxypyrimidinyl)oxy)phenyl)urea 8 4-fluoro-3- C: 52.5N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′- 480 trifluoromethyl H: 3.67(4-(2-(N-methylaminoformoxyl)-5-(4- N: 14.5methoxypyrimidinyl)oxy)phenyl)urea 9 4-bromo-3- C: 46.8N-(4-bromo-3-(trifluoromethyl)phenyl)-N′- 541 trifluoromethyl H: 3.00(4-(2-(N-methylaminoformoxyl)-5-(4- N: 13.1methoxypyrimidinyl)oxy)phenyl)urea

TABLE 4 substituted isoquinoline derivatives

Compound Elementary Mass No. R₃ analysis Name spectrum m/e 104-chloro-3- C: 56.5 N-(4-chloro-3-(trifluoromethyl)phenyl)-N′- 530.5trifluoromethyl H: 3.70 (4-(2-(N-methylaminoformoxyl)- N: 13.24-(7-amino-isoquinolyl)oxy)phenyl)urea 11 4-fluoro-3- C: 58.5N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′- 514 trifluoromethyl H: 3.81(4-(2-(N-methylaminoformoxyl)- N: 13.84-(7-amino-isoquinolyl)oxy)phenyl)urea 12 4-bromo-3- C: 52.4N-(4-bromo-3-(trifluoromethyl)phenyl)-N′- 574 trifluoromethyl H: 3.44(4-(2-(N-methylaminoformoxyl)- N: 12.44-(7-amino-isoquinolyl)oxy)phenyl)urea

TABLE 5 substituted pyrrole derivatives

Compound Elementary Mass No. R₃ analysis Name spectrum m/e 134-chloro-3- C: 54.1 N-(4-chloro-3-(trifluoromethyl)phenyl)-N′- 467.5trifluoromethyl H: 4.01 (4-(2-(N-methylaminoformoxyl)-4-(5- N: 12.2methyl-pyrryl)oxy)phenyl)urea 14 4-fluoro-3- C: 55.8N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′- 451 trifluoromethyl H: 4.02(4-(2-(N-methylaminoformoxyl)-4-(5- N: 12.6methyl-pyrryl)oxy)phenyl)urea 15 4-bromo-3- C: 49.5N-(4-bromo-3-(trifluoromethyl)phenyl)-N′- 511 trifluoromethyl H: 3.70(4-(2-(N-methylaminoformoxyl)-4-(5- N: 11.0methyl-pyrryl)oxy)phenyl)urea

TABLE 6 substituted quinoline derivatives

Compound Elementary Mass No. R₃ analysis Name spectrum m/e 164-chloro-3- C: 59.2 N-(4-chloro-3-(trifluoromethyl)phenyl)-N′- 488.5trifluoromethyl H: 3.70 (4-(4-(6-methoxyquinolinyl)oxy))phenyl)urea N:8.77 17 4-fluoro-3- C: 61.3 N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-472 trifluoromethyl H: 3.81 (4-(4-(6-methoxyquinolinyl)oxy))phenyl)ureaN: 8.80 18 4-bromo-3- C: 54.0 N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-532 trifluoromethyl H: 3.40 (4-(4-(6-methoxyquinolinyl)oxy))phenyl)ureaN: 7.79

TABLE 7 substituted quinazoline derivatives

Compound Elementary Mass No. R₃ analysis Name spectrum m/e 194-chloro-3- C: 55.5 N-(4-chloro-3-(trifluoromethyl)phenyl)-N′- 477.5trifluoromethyl H: 2.91 (4-(4-(7 fluoroquinazolinyl)oxy)phenyl)urea N:11.6 20 4-fluoro-3- C: 57.7 N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-461 trifluoromethyl H: 2.75 (4-(4-(7 fluoroquinazolinyl)oxy)phenyl)ureaN: 12.2 21 4-bromo-3- C: 50.5 N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-522 trifluoromethyl H: 2.71 (4-(4-(7 fluoroquinazolinyl)oxy)phenyl)ureaN: 10.8

TABLE 8 substituted pyrimidine derivatives

Compound Elementary Mass No. R₃ analysis Name spectrum m/e 224-chloro-3- C: 52.1 N-(4-chloro-3-(trifluoromethyl)phenyl)-N′- 439.5trifluoromethyl H: 3.30 (4-(5-(4-methoxypyrimidinyl)oxy)phenyl) N: 12.823 4-fluoro-3- C: 54.2 N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′- 423trifluoromethyl H: 3.38 (4-(5-(4-methoxypyrimidinyl)oxy)phenyl) N: 13.324 4-bromo-3- C: 47.2 N-(4-bromo-3-(trifluoromethyl)phenyl)-N′- 484trifluoromethyl H: 2.99 (4-(5-(4-methoxypyrimidinyl)oxy)phenyl) N: 11.4

TABLE 9 substituted isoquinoline derivatives

Compound Elementary Mass No. R₃ analysis % Name spectrum m/e 254-chloro-3- C: 58.4 N-(4-chloro-3-(trifluoromethyl)phenyl)-N′- 473.5trifluoromethyl H: 3.38 (4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea N:11.81 26 4-fluoro-3- C: 60.7 N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-457 trifluoromethyl H: 3.70 (4-(4-(7-amino-isoquinolyl)oxy)phenyl)ureaN: 13.5 27 4-bromo-3- C: 53.5 N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-518 trifluoromethyl H: 3.40 (4-(4-(7-amino-isoquinolyl)oxy)phenyl)ureaN: 10.7

TABLE 10 substituted pyrrole derivatives

Compound Elementary Mass No. R₃ analysis % Name spectrum m/e 284-chloro-3- C: 55.7 N-(4-chloro-3-(trifluoromethyl)phenyl)-N′- 410.5trifluoromethyl H: 3.75 (4-(4-(2-methyl-pyrryl)oxy)phenyl)urea N: 10.429 4-fluoro-3- C: 58.2 N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′- 395trifluoromethyl H: 4.01 (4-(4-(2-methyl-pyrryl)oxy)phenyl)urea N %: 10.530 4-bromo-3- C: 50.3 N-(4-bromo-3-(trifluoromethyl)phenyl)-N′- 455trifluoromethyl H: 3.49 (4-(4-(2-methyl-pyrryl)oxy)phenyl)urea N: 9.41

TABLE 11 polysubstituted quinoline derivatives

Compound Elementary Mass No. R₃ analysis Name spectrum m/e 314-chloro-3- C: 64.6 N-(4-chloro-3-ethylphenyl)-N′-(4-(4-(6- 466.5 ethylH: 4.71 methoxyl-7-fluoro-quinolinyl)oxy)phenyl)urea N: 9.20 324-ethyl-3- C: 62.7 N-(4-ethyl-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-501.5 trifluoromethyl H: 4.36methoxyl-7-fluoro-quinolinyl)oxy)phenyl)urea N: 8.22

TABLE 12 polysubstituted quinazoline derivatives

Compound Elementary Mass No. R₃ analysis Name spectrum m/e 334-chloro-3- C: 64.1 N-(4-chloro-3-ethylphenyl)-N′-(4-(4-(6- 451.5 ethylH: 4.70 methyl-7-fluoro-quinolinyl)oxy)phenyl)urea N: 12.3 34 4-ethyl-3-C: 61.8 N-(4-ethyl-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6- 485trifluoromethyl H: 4.01 methyl-7-fluoro-quinolinyl)oxy)phenyl)urea N:11.38

TABLE 13 substituted quinoline derivatives

Compound Elementary Mass No. R₂ analysis Name spectrum m/e 35 2-methylC: 59.1 N-(4-chloro-3-methoxyphenyl)-N′- 550.5 H: 4.32(2-methyl-4-(2-(N-ethylcarbamyl)-4- N: 12.5(7-nitryl-quinolinyl)oxy)phenyl)urea 36 2-methoxy C: 57.2N-(4-chloro-3-methoxyphenyl)-N′- 566.5 H: 4.36(2-methoxyl-4-(2-(N-ethylcarbamyl)-4- N: 12.2(7-nitryl-quinolinyl)oxy)phenyl)urea

TABLE 14 substituted quinazoline derivatives

Compound Elementary Mass No. R₂ analysis Name spectrum m/e 37 2-fluoroC: 56.1 N-(4-chloro-3-methoxyphenyl)-N′- 579.5 H: 4.17(2-fluoro-4-(2-(N-propylcarbamyl)-4- N: 9.72(7-trifluoromethylquinolinyl)oxy))phenyl)urea 38 2-trifluoro- C: 53.4N-(4-chloro-3-methoxyphenyl)-N′- 629.5 methyl H: 3.91(2-trifluoromethyl-4-(2-(N-propylcarbamyl)-4- N: 8.58(7-trifluoromethylquinolinyl)oxy))phenyl)urea

Determination of Antitumor Activity

1. Inhibitory activity of the compound of the present invention on rafkinase

[Test Method]

Raf-1 inhibitor screening by chemoluminescence method

[Instruments]

Westernblot electrophoresis apparatus Rotaryshaker

[Test Materials]

Raf-1(truncated), Magnesium/ATP Cocktail, MEK1 unactive

[Tested Samples]

Compounds 1-38

[Positive Control]

Sorafenib

${{Inhibiton}\mspace{14mu} {rate}\mspace{11mu} \%} = {\frac{\begin{matrix}{{{Gray}\mspace{14mu} {value}\mspace{14mu} {of}\mspace{14mu} {the}\mspace{14mu} {negative}\mspace{14mu} {control}{\mspace{11mu} \;}{group}} -} \\{{gray}\mspace{14mu} {value}\mspace{14mu} {of}\mspace{14mu} {the}{\mspace{11mu} \;}{drug}\text{-}{treated}\mspace{14mu} {group}}\end{matrix}}{{Gray}\mspace{14mu} {value}\mspace{14mu} {of}\mspace{14mu} {the}\mspace{14mu} {negative}\mspace{14mu} {control}\mspace{14mu} {group}} \times 100\%}$

[Results]

TABLE 13 Inhibition of compounds 1-16 and positive control medicine onraf kinase Compound Final concentration Inhibition Compound Finalconcentration Inhibition No. 1.0*10⁻⁵ mol/ml rate % Activity No.1.0*10⁻⁵ mol/ml rate % Activity 1 1 75.0 + 9 1 99.2 + 2 1 61.1 + 10 13.5 3 1 56.3 + 11 1 11.2 4 1 82.2 + 12 1 12.1 5 1 98.9 + 13 1 55.0 + 6 180.1 + 14 1 41.3 7 1 99.1 + 15 1 35.5 8 1 44..5 16 1 62.1 + Positive 185.7 + control medicine

TABLE 14 Inhibition of compounds 17-38 and positive control medicine onraf kinase Compound Final concentration Inhibition Compound Finalconcentration Inhibition No. 1.0*10⁻⁵ mol/ml rate % Activity No.1.0*10⁻⁵ mol/ml rate % Activity 17 1 58.2 + 28 1 33.5 18 1 34.4 29 185.3 + 19 1 93.5 + 30 1 16.8 20 1 87.7 + 31 1 89.4 + 21 1 98.9 + 32 190.5 + 22 1 88.1 + 33 1 92.3 + 23 1 89.9 + 34 1 96.2 + 24 1 91.3 + 35 145.3 25 1 11.5 36 1 81.2 + 26 1 15.3 37 1 81.5 + 27 1 8.8 38 1 88.1 +Positive 1 85.7 + control medicine

The test results of inhibitory activity of the compound on raf kinaseshowed that the inhibitory activity of the compound in the presentinvention is better than or equivalent to positive control medicinesorafenib. The test results indicate that these compounds can affect thesurvival, proliferation and disease progression of tumor cells throughinhibiting the raf kinase and blocking the ras protein signaltransduction cascade of tumor cells. The compound of the presentinvention has potential of being applied to treat tumor and leukemia.

2. Experimental therapeutic action of the compound in the presentinvention on S180 sarcoma mice

[Test Materials]

Test animals: ICR mice, 18-25 g

Tumor types: mice S180 sarcoma, provided by Shanghai Institute ofMateria Medica, Chinese Academy of Sciences.

-   -   Positive control medicine: Sorafenib    -   Tested samples: compounds 1-38

[Test Method]

18-25 g female ICR mice and well grown 7-11 day old mice sarcoma S180tumor seeds were selected, and the seeds were inoculated into thesubcutaneous at the right axillary. After inoculated 24 hours, thesemice were randomly divided into cages and orally administrated 60 mg/kgfor 9 days. On 10 day, the animals were killed and weighed, and thetumor weights were weighed to calculate average tumor weight in eachgroup, followed by calculating the tumor inhibition rate according tothe following formula and T test.

${{Tumor}\mspace{14mu} {growth}\mspace{14mu} {inhibition}\mspace{14mu} {rate}} = {\frac{\begin{matrix}{{{Average}\mspace{14mu} {tumor}\mspace{14mu} {weight}{\mspace{11mu} \;}{in}\mspace{14mu} {the}\mspace{14mu} {control}\mspace{14mu} {group}} -} \\{{average}\mspace{14mu} {tumor}\mspace{14mu} {weight}\mspace{14mu} {in}\mspace{14mu} {the}\mspace{14mu} {treatment}\mspace{14mu} {group}}\end{matrix}}{{average}\mspace{14mu} {tumor}\mspace{14mu} {weight}{\mspace{11mu} \;}{in}\mspace{14mu} {the}\mspace{14mu} {treatment}\mspace{14mu} {group}} \times 100\%}$

[Determination Results]

TABLE 15 Tumor growth inhibition rate of compounds 1-3 and sorafenib onmice S180 sarcoma Administration Animal number Weight (g) Tumor weightInhibition Groups Dosage methods Start Final Start Final x ± SD(g) rate(%) P value Normal 0.4 ml/mouse ig 20 20 18.9 ± 1.5 22.0 ± 3.4 1.61 ±0.36 saline Sora 60 mg/kg ig 10 10 18.8 ± 1.2 21.7 ± 2.4 0.71 ± 0.3055.9 <0.05 Compound 1 60 mg/kg ig 10 10 18.7 ± 1.9 22.3 ± 1.3 0.99 ±0.20 38.5 <0.05 Compound 2 60 mg/kg ig 10 10 18.9 ± 1.7 20.9 ± 2.3 0.87± 0.24 46.0 <0.05 Compound 3 60 mg/kg ig 10 10 18.0 ± 1.1 20.2 ± 2.50.75 ± 0.36 53.4 <0.05

TABLE 16 Tumor growth inhibition rate of compounds 1-12 and sorafenib onmice S180 sarcoma (%) 1 2 3 4 5 6 7 8 9 10 11 12 Sorafenib Mice 38.524.1 53.4 50.1 49.3 51.2 21.5 58.2 55.9 55.2 54.2 45.7 55.9% S180sarcoma

TABLE 17 Tumor growth inhibition rate of compounds 13-26 and sorafenibon mice S180 sarcoma (%) 13 14 15 16 17 18 19 20 21 22 23 24 25 26 Mice21.2 18.2 56.7 33.6 44.2 35.7 50.8 54.6 59.7 52.1 51.5 54.6 55.8 7.6S180 sarcoma

TABLE 18 Tumor growth inhibition rate of compounds 27-38 and sorafenibon mice S180 sarcoma (%) 27 28 29 30 31 32 33 34 35 36 37 38 Mice 55.111.5 14.2 55.2 50.3 49.8 55.3 52.6 55.8 55.1 24.1 59.2 S180 sarcoma

3. Experimental therapeutic action of the compound in the presentinvention on Human colon cancer HT-29 transplantable tumor in nude mice.

[Test Materials]

-   -   Test animals: Female BALB/cA nude mice, 35-40 day old, with        weight of 18-22 g.    -   Tumor seeds: Human colon cancer HT-29 transplantable tumor in        nude mice, established by inoculating human colon cancer HT-29        cell strains subcutaneously in nude mice    -   Positive control medicine: Sorafenib    -   Tested samples: Compounds 1-38

[Test Method]

Take eugenic tumor tissues and cut into about 1.5 mm³, and thenincoculate subcutaneously at the right armpit of nude mice under thesterile conditions. The diameter of the transplantable tumor in nudemice was determined with a vernier caliper, and the animals were dividedinto groups after the tumors were grown to 100-300 mm³. Using the methodof measuring the tumor diameter, dynamically observe the antitumoreffects of tested materials. The diameter of the tumor was determinedthree times every week and the mouse weight was weighed at the sametime. The mice were intragastrically administrated with Sorafenib andtested drugs, 60 mg/kg, for continuous 9 times. The solvent wasintragastrically administrated as the control for continuous 9 times.Equal amount of control was administrated in the negative control group.Tumor volume (TV) is calculated as: TV=1/2×a×b², wherein a and brespectively represent length and width.

Relative tumor volume (RTV) is calculated as: RTV=TV_(t)/TV₀, whereinTV₀ is the tumor volume when administrated according to different cagesand TV_(t) is the tumor volume measured each time.

Relative tumor reproduction rate T/C (%) is calculated as follows:

${T\text{/}{C(\%)}} = {\frac{T_{RTV}}{C_{RTV}} \times 100}$

T_(RTV): RTV in the treatment group; C_(RTV): RTV in the negativecontrol group.

The test results used relative tumor reproduction rate T/C (%) asevaluating indicator of anti-tumor activity.

Evaluation of in vivo anti-tumor activity

T/C % Evaluation ≧60 (−) No activity 60-50 (+/−) Marginal activity 50-40(+) Moderate-strength activity 40-10 (++) High-strength activity ≦10(+++)Extremenly high-strength activity

[Determination results]

TABLE 19 Experimental treatment of compounds 1-3 and Sorafenib on humancolon cancer HT-29 transplantable tumor in nude mice Animal Dosagenumber Weight (g) TV T/C Groups mg/kg Start Final d0 d13 d0 d13 RTV (%)Control 6 6 18.8 ± 1.1 19.6 ± 0.9 133 ± 60 626 ± 226 5.07 ± 1.39 Solventcontrol 6 6 19.7 ± 0.6 20.8 ± 0.8 133 ± 32 547 ± 172 4.15 ± 0.93 81.85Sorafenib 60 6 6 19.9 ± 1.1 20.4 ± 1.4 133 ± 33 308 ± 86  2.36 ± 0.5746.48** Compound 1 60 6 6 19.5 ± 1.0 20.5 ± 1.1 128 ± 34 359 ± 108 2.81± 1.05 55.42** Compound 2 60 6 6 19.9 ± 0.8 21.0 ± 0.8 133 ± 23 265 ±100 2.00 ± 0.41 39.45** Compound 3 60 6 6 19.1 ± 1.0 19.7 ± 1.4 133 ± 18322 ± 129 2.40 ± 0.67 47.34**

TABLE 20 Relative tumor reproduction rate of compounds 1-10 andSorafenib on Human colon cancer HT-29 transplantable tumor in nude miceT/C (%) 1 2 3 4 5 6 7 8 9 10 Sorafenib Human colon cancer 55.42 39.4547.34 33.15 38.24 39.58 40.1 37.6 35.2 78.9 46.48 HT-29 transplantabletumor in nude mice

TABLE 21 Relative tumor reproduction rate of compounds 11-20 andSorafenib on human colon cancer HT-29 transplantable tumor in nude miceT/C (%) 11 12 13 14 15 16 17 18 19 20 Human colon 81.2 80.5 81.2 40.185.2 55.2 54.7 40.2 39.4 40.1 cancer HT-29 transplantable tumor in nudemice

TABLE 22 Relative tumor reproduction rate of compounds 21-30 andSorafenib on human colon cancer HT-29 transplantable tumor in nude miceT/C (%) 21 22 23 24 25 26 27 28 29 30 Human colon 38.7 41.2 39.7 37.879.5 41.1 80.3 39.1 78.3 79.5 cancer HT-29 transplatable tumor in nudemice

TABLE 23 Relative tumor reproduction rate of compounds 31-38 andSorafenib on human colon cancer HT-29 transplantable tumor in nude miceT/C (%) 31 32 33 34 35 36 37 38 Human colon 50.3 46.2 39.9 45.9 51.241.5 39.5 41.2 cancer HT-29 transplantable tumor in nude mice

The results of the above in vivo and vitro tumor inhibition tests showedthat the inhibiting effects of such derivatives on S180 sarcoma in miceand human colon cancer HT-29 transplantable tumor in nude mice werebetter than or equivalent to positive control medicine sorafenib.Thetest results showed that the compound of the present invention or thepharmaceutically acceptable salt thereof can be used for treating tumoror leukemia. The pharmacodynamic experiments of the compounds in thepresent invention, positive control medicine sorafenib and compounds A′,B′ and C′ with no substituent or only amino formyl in A ring on humanlung cancer cell strain A549, human high-metastic lung cancer cellstrain 95D, lung cancer cell A549, human umbilical vein endothelial cellHUVEC cell growth and lumen formation, human lung cancer A549 celltransplantation tumor model in nude mice, human liver cancer cellbel-7402 transplantation tumor model in nude mice, and renal carcinomacell line GCR-1 transplantation tumor model in nude mice were carriedout to verify the effect of the compounds of the present invention.

Sorafenib was abbreviated as Sorafenib hereafter, and the compounds A′,B′ and C′ were respectively prepared according the method ofCN200810129360.6, which were compounds with no substituent in A ring,wherein A′ is

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-quinolinyl)oxy)phenyl)urea,B′is

N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-pyrimidinyl)oxy)phenyl)urea,and C′

N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-pyrryl)oxy)phenyl)urea.

4. Using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromideassay (MTT) to detect the inhibition effects on the growth of human lungcancer cell strain A549

[Test materials] MTT working solution, F12 medium containing 10% FBS,continuous injectors

[Tested compound] Compounds 1-38

[Positive control medicine] Sorafenib, compounds A′, B′, C′ with nosubstituent or only amino formyl in A ring

The inhibition rate is calculated as follows:

${{Cell}\mspace{14mu} {reproduction}\mspace{14mu} {rate}\mspace{11mu} \%} = {1 - {\frac{\begin{pmatrix}{{{Relative}\mspace{14mu} {OD}\mspace{14mu} {value}\mspace{14mu} {of}\mspace{14mu} {conrol}\mspace{14mu} {well}} -} \\{{Relative}\mspace{14mu} {OD}\mspace{14mu} {value}\mspace{14mu} {of}\mspace{14mu} {drug}\mspace{14mu} {well}}\end{pmatrix}}{{Relative}\mspace{14mu} {OD}\mspace{14mu} {value}\mspace{14mu} {of}\mspace{14mu} {conrol}\mspace{14mu} {well}} \times 100\%}}$

Relative OD value of conrol well=OD value of control well−OD value ofblank well

Relative OD value of drug well=OD value of drug well−OD value of blankwell

[Screening Results]

TABLE 24 Inhibition effects of compounds 1-18 on the growth of humanlung cancer cell A549 Compound Final concentration Inhibition No.1.0*10⁽⁻⁵⁾ mol/L rate Activity 1 1 50.20% + 2 1 61.11% ++ 3 1 69.67% ++4 1 91.12% +++ 5 1 93.50% +++ 6 1 92.50% +++ 7 1 45.30% + 8 1 60.67% ++9 1 61.54% ++ 10 1 74.50% ++ 11 1 69.20% ++ 12 1 42.31% + 13 1 64.23% ++14 1 67.25% ++ 15 1 72.17% ++ 16 1 89.71% +++ 17 1 88.52% +++ 18 190.71% +++

TABLE 25 Inhibition effects of compounds 19-38 on the growth of humanlung cancer cell A549 Compound Final concentration Inhibition No.1.0*10⁽⁻⁵⁾ mol/ml rate Activity 19 1 91.23% +++ 20 1 88.32% +++ 21 189.21% +++ 22 1 82.49% +++ 23 1 55.41% + 24 1 86.32% +++ 25 1 63.26% +26 1 55.50% + 27 1 64.71% + 28 1 88.52% +++ 29 1 86.42% +++ 30 1 85.47%+++ 31 1 46.78% + 32 1 45.76% + 33 1 57.53% + 34 1 59.52% + 35 155.74% + 36 1 72.45% ++ 37 1 71.65% ++ 38 1 74.56% ++ Posive Sorafenib 154.60% + Compound A′ 1 58.51% + Compound B′ 1 61.62% + Compound C′ 162.25% +

5 Inhibition effects of compounds on human high-metastic lung cancercell 95D migration

[Test materials] Boyden Chamber Transwell chamber (with pore size of 8μm), human high-metastic lung cancer cell 95D cell strain, 1640 mediumcontaining 10% FBS, 1640 medium containing no serum

[Tested compound] Compounds 1-38

[Positive control medicine] Sorafenib, compounds A′, B′, C′ with nosubstituent or only amino formyl in A ring

The inhibition rate is calculated as follows:

${{Cell}\mspace{14mu} {migration}\mspace{14mu} {inhibition}\mspace{14mu} {rate}\mspace{11mu} \%} = {\frac{\begin{pmatrix}{{{migrated}\mspace{14mu} {cell}\mspace{14mu} {number}{\mspace{11mu} \;}{in}\mspace{14mu} {the}\mspace{14mu} {chamber}\mspace{14mu} {containing}\mspace{14mu} {no}\mspace{14mu} {drug}} -} \\{{migrated}\mspace{14mu} {cell}\mspace{14mu} {number}{\mspace{11mu} \;}{in}\mspace{14mu} {the}\mspace{14mu} {chamber}\mspace{14mu} {containing}\mspace{14mu} {drug}}\end{pmatrix}}{{migrated}\mspace{14mu} {cell}\mspace{14mu} {number}\mspace{14mu} {in}\mspace{14mu} {the}\mspace{14mu} {chamber}\mspace{14mu} {containing}\mspace{14mu} {no}\mspace{14mu} {drug}} \times 100\%}$

[Screening Results]

TABLE 26 Inhibition effects of compounds 1-18 on human high- metasticlung cancer 95D cell strain migration Compound Final concentrationInhibition No. 1.0*10⁽⁻⁵⁾ mol/L rate Activity 1 1 80.21% ++ 2 1 85.17%+++ 3 1 96.64% +++ 4 1 96.38% +++ 5 1 97.51% +++ 6 1 93.71% +++ 7 189.34% +++ 8 1 89.56% +++ 9 1 91.42% +++ 10 1 61.43% + 11 1 78.66% ++ 121 66.79% ++ 13 1 65.45% ++ 14 1 57.57% + 15 1 63.68% + 16 1 89.31% +++17 1 90.52% +++ 18 1 93.73% +++

TABLE 27 Inhibition effects of compounds 19-38 on human high- metasticlung cancer 95D cell strain migration Compound Final concentrationInhibition No. 1.0*10⁽⁻⁵⁾ mol/L rate Activity 19 1 90.21% +++ 20 189.12% +++ 21 1 88.76% +++ 22 1 88.77% +++ 23 1 85.53% +++ 24 1 85.48%+++ 25 1 59.76% + 26 1 60.52% + 27 1 59.77% + 28 1 70.53% + 29 161.44% + 30 1 69.62% ++ 31 1 76.18% ++ 32 1 66.92% ++ 33 1 77.52% ++ 341 63.65% + 35 1 68.47% ++ 36 1 84.59% +++ 37 1 79.25% + 38 1 80.53% +Positive Sorafenib 1 62.32% + Compound A′ 1 63.51% + Compound B′ 161.60% + Compound C′ 1 63.20% ++

6. Effects of tested compounds on the adhesive ability of lung cancercell A549

[Test materials] gelatin, CCK8, poly-lysine (PLL), A549 cell stains,1640 medium containing 10% FBS

[Tested compound] Compounds 1-38 to be tested

[Positive control medicine] Sorafenib, compounds A′, B′, C′ with nosubstituent or only amino formyl in A ring

[Screening Results]

The inhibition rate is calculated as follows:

${{Inhibition}\mspace{14mu} {rate}\mspace{14mu} {of}\mspace{14mu} {cell}\mspace{14mu} {adhesion}\mspace{14mu} \%} = {\frac{\begin{matrix}{{{Cell}\mspace{14mu} {group}\mspace{14mu} {without}{\mspace{11mu} \;}{treatment}}\;} \\{\; {( {{glutin}\mspace{14mu} {adhesion}\mspace{14mu} {{OD}/{PLL}}{\mspace{11mu} \;}{adhesion}\mspace{14mu} {OD}\mspace{14mu} {value}}\; ) -}} \\{{dosing}\mspace{14mu} {cell}\mspace{14mu} {group}\mspace{11mu} ( {{glutin}\mspace{14mu} {adhesion}\mspace{14mu} {{OD}/{PLL}}\mspace{14mu} {adhesion}\mspace{14mu} {OD}\mspace{14mu} {value}} )}\end{matrix}}{\begin{matrix}{{Cell}\mspace{14mu} {group}\mspace{14mu} {without}\mspace{14mu} {treatment}} \\( {{glutin}\mspace{14mu} {adhesion}\mspace{14mu} {{OD}/{PLL}}\mspace{14mu} {adhesion}\mspace{14mu} {OD}\mspace{14mu} {value}} )\end{matrix}} \times 100\%}$

Dosing dosing cells without cell group (gelatin adhesive OD/PLL adhesionOD)−Dosing cell group (gelatin adhesive OD/PLL adhesion OD value)

Dosing dosing cells without cell group (gelatin adhesive OD/PLL adhesionOD value)

TABLE 28 Inhibition effects of compounds 1-18 on the adhesion ability ofhuman lung cancer cell A549 Compound Final concentration Inhibition No.1.0*10⁽⁻⁵⁾ mol/L rate Activity 1 1 60.22% + 2 1 75.15% ++ 3 1 79.66% ++4 1 89.71% +++ 5 1 87.58% +++ 6 1 93.59% +++ 7 1 68.34% ++ 8 1 61.56% +9 1 85.32% +++ 10 1 64.57% + 11 1 59.63% + 12 1 62.30% + 13 1 63.39% +14 1 67.51% ++ 15 1 68.63% ++ 16 1 90.77% +++ 17 1 97.50% +++ 18 193.72% +++

TABLE 29 Inhibition effects of compounds 19-38 on the adhesion abilityof human lung cancer cell A549 Compound Final concentration InhibitionNo. 1.0*10⁽⁻⁵⁾ mol/L rate Activity 19 1 90.28% +++ 20 1 95.31% +++ 21 190.22% +++ 22 1 72.44% ++ 23 1 77.56% ++ 24 1 73.30% ++ 25 1 63.51% + 261 68.57% ++ 27 1 94.77% +++ 28 1 90.53% +++ 29 1 91.40% +++ 30 1 92.44%+++ 31 1 66.77% ++ 32 1 64.73% + 33 1 77.59% ++ 34 1 76.54% ++ 35 175.72% ++ 36 1 74.50% ++ 37 1 71.74% ++ 38 1 75.53% ++ Positive medicine1 72.66% ++ Sorafenib Compound A′ 1 71.55% ++ Compound B′ 1 69.26% ++Compound C′ 1 68.62% ++

7. Effects of tested compounds on the growth of human umbilical veinendothelial cell HUVEC cell by CCK8 method

[Test materials] CCK8, human umbilical vein endothelial cell HUVEC cell,1640 medium containing 10% FBS

[Tested compound] Compounds 1-38 to be tested

[Positive control medicine] Sorafenib, compounds A′, B′, C′ with nosubstituent or only amino formyl in A ring

[Screening Results]

${{Cell}\mspace{14mu} {reproduction}\mspace{14mu} {rate}\mspace{11mu} \%} = {1 - {\frac{\begin{pmatrix}{{{Relative}\mspace{14mu} {OD}\mspace{14mu} {value}\mspace{14mu} {of}\mspace{14mu} {conrol}\mspace{14mu} {well}} -} \\{{Relative}\mspace{14mu} {OD}\mspace{14mu} {value}\mspace{14mu} {of}\mspace{14mu} {drug}\mspace{14mu} {well}}\end{pmatrix}}{{Relative}\mspace{14mu} {OD}\mspace{14mu} {value}\mspace{14mu} {of}\mspace{14mu} {conrol}\mspace{14mu} {well}} \times 100\%}}$

Relative OD value of conrol well=OD value of control well−OD value ofblank well

Relative OD value of drug well=OD value of drug well−OD value of blankwell

TABLE 30 Inhibition effects of compounds 1-18 on the growth of humanumbilical vein endothelial cell HUVEC cell Compound Final concentrationInhibition No. 1.0*10⁽⁻⁵⁾ mol/L rate Activity 1 1 9.20% 2 1 13.11% 3 127.67% 4 1 5.12% 5 1 8.50% 6 1 7.50% 7 1 8.30% 8 1 13.67% 9 1 11.54% 101 14.50% 11 1 29.20% 12 1 12.31% 13 1 24.23% 14 1 17.25% 15 1 38.17% 161 10.71% 17 1 13.52% 18 1 5.71%

TABLE 31 Inhibition effects of compounds 19-38 on the growth of humanumbilical vein endothelial cell HUVEC cell Compound Final concentrationInhibition No. 1.0*10⁽⁻⁵⁾ mol/L rate Activity 19 1 5.23% 20 1 7.32% 21 111.21% 22 1 12.49% 23 1 17.41% 24 1 13.32% 25 1 23.26% 26 1 32.50% 27 124.73% 28 1 20.55% 29 1 21.40% 30 1 25.46% 31 1 26.70% 32 1 24.77% 33 117.50% 34 1 19.52% 35 1 5.74% 36 1 12.45% 37 1 19.65% 38 1 14.56%Positive medicine 1 22.61% Sorafenib Compound A′ 1 23.11% Compound B′ 131.64% Compound C′ 1 22.27%

8. Inhibition effects of compounds on the lumen formation ability ofhuman umbilical vein endothelial cell HUVEC

[Experimental principles] The human umbilical vein endothelial cellshave ability of spontaneously forming blood lumen on Matrigel, which canbe used to simulate the process of angiogenesis in vivo. We usedMatrigel method to investigate the effects of the compound on the lumenformation ability of human umbilical vein endothelial cell HUVEC.

[Test materials] HUVEC (taking generation 3 to 5 cells for experimentsafter obtained from primary separation and cultured at 37 under theconditions of 5% CO₂), Matrigel, cell culture medium M199.

[Tested compound] Compounds 1-38

[Positive control medicine] Sorafenib, compounds A′, B′, C′ with nosubstituent or only amino formyl in A ring

[Screening Results]

The inhibition rate is calculated as follows:

${{Lumen}\mspace{14mu} {formation}\mspace{14mu} {inhibition}\mspace{14mu} {rate}\mspace{11mu} \%} = {\frac{\begin{pmatrix}{{{length}\mspace{14mu} {sum}\mspace{14mu} {of}\mspace{14mu} {lumen}\mspace{14mu} {without}\mspace{14mu} {dosing}} -} \\{{length}\mspace{14mu} {sum}\mspace{14mu} {of}\mspace{14mu} {lumen}\mspace{14mu} {after}{\mspace{11mu} \;}{dosing}}\end{pmatrix}}{{length}\mspace{14mu} {sum}\mspace{14mu} {of}\mspace{14mu} {lumen}\mspace{14mu} {without}\mspace{14mu} {dosing}} \times 100\%}$

TABLE 32 Inhibition effects of compounds 1-18 on the lumen formationability of human umbilical vein endothelial cell HUVEC Compound Finalconcentration Inhibition No. 1.0*10⁽⁻⁵⁾ mol/L rate Activity 1 1 72.22%++ 2 1 75.14% ++ 3 1 76.61% ++ 4 1 86.77% +++ 5 1 87.50% +++ 6 1 90.55%+++ 7 1 60.34% + 8 1 78.59% ++ 9 1 91.45% +++ 10 1 50.34% + 11 1 79.63%++ 12 1 52.37% + 13 1 53.36% + 14 1 67.53% ++ 15 1 68.64% ++ 16 1 80.79%++ 17 1 67.50% ++ 18 1 63.71% +

TABLE 33 Inhibition effects of compounds 19-38 on the lumen formationability of human umbilical vein endothelial cell HUVEC Compound Finalconcentration Inhibition No. 1.0*10⁽⁻⁵⁾ mol/L rate Activity 19 1 89.22%+++ 20 1 88.54% +++ 21 1 89.47% +++ 22 1 86.72% +++ 23 1 97.56% +++ 24 193.78% +++ 25 1 79.79% ++ 26 1 70.53% ++ 27 1 89.75% +++ 28 1 89.52% +++29 1 86.43% +++ 30 1 90.66% +++ 31 1 66.77% ++ 32 1 54.74% + 33 1 67.51%++ 34 1 66.52% ++ 35 1 65.70% ++ 36 1 64.50% + 37 1 71.72% ++ 38 165.55% ++ Positive medicine 1 55.60% ++ Sorafenib Compound A′ 1 60.51%++ Compound B′ 1 61.62% ++ Compound C′ 1 60.50% ++

9. Tumor inhibition rate of tested compounds on human lung cancer A549cell transplantation model in nude mice

[Test animals] female BALB/cA nude mice, 35-40 days old, with weight of18-22g. There were 12 mice in the negative conrol group and 6 mice inthe treatment group.

[Test method] Take eugenic tumor tissues and cut into about 1.5 mm³, andthen incoculate subcutaneously at the right armpit of nude mice underthe sterile conditions. The diameter of the transplantable tumor in nudemice was determined with a vernier caliper, and the animals were dividedinto groups after the tumors were grown to 100-300 mm³.Using the methodof measuring the tumor diameter, dynamically observe the antitumoreffects of tested materials.

The diameter of the tumor was determined three times every week and themouse weight was weighed at the same time. The dosage of the compoundwas 60 mg/kg, 6 times every week for 3 weeks. Sorafenib was oraladministrated with dosage of 60 mg/kg, 6 times every week for 3 weeks.Equal amount of normal saline was administrated in the negative controlgroup. Observe for one week after administration.

[Detection Indicators and Calculation Methods]

(1) Tumor volume (TV) is calculated as:

-   -   TV=1/2×a×b²    -   wherein a and b respectively represents length and width.

(2) Relative tumor volume (RTV) is calculated as:

-   -   RTV=TV_(t)/TV_(0°)

wherein TV₀ is the tumor volume when administrated according todifferent cages and TV_(t) is the tumor volume measured each time.

(3) Relative tumor reproduction rate T/C (%) is calculated as follows:

${T\text{/}{C(\%)}} = {\frac{T_{RTV}}{C_{RTV}} \times 100}$

T_(RTV): RTV in the treatment group; C_(RTV): RTV in the negativecontrol group. The test results used relative tumor reproduction rateT/C (%) as evaluating indicator of anti-tumor activity.

[Screening results] There was no mortality for the animals in the groupof compounds and Sorafenib in the experiments with less toxicity.

TABLE 34 Relative tumor reproduction rate of compounds and Sorafenib onhuman lung cancer A549 transplantation tumor in nude mice T/C (%)Relative tumor reproduction rate of compounds 1-10 and Sorafenib onhuman lung cancer A549 transplantation tumor in nude mice T/C (%) 1 2 34 5 6 7 8 9 10 human lung 38.7 35.3 37.2 12.3 11.6 15.9 39.3 30.5 38.730.1 cancer A549 transplantation tumor Relative tumor reproduction rateof compounds 11-20 and Sorafenib on human lung cancer A549transplantation tumor in nude mice T/C (%) 11 12 13 14 15 16 17 18 19 20human lung 43.7 36.3 38.4 32.2 38.7 9.9 7.3 12.5 7.7 12.1 cancer A549transplantation tumor Relative tumor reproduction rate of compounds21-30 and Sorafenib on human lung cancer A549 transplantation tumor innude mice T/C (%) 21 22 23 24 25 26 27 28 29 30 human lung 13.7 15.317.1 15.6 39.4 33.1 36.3 13.6 14.7 13.1 cancer A549 transplantationtumor Relative tumor reproduction rate of compounds 31-38 and Sorafenibon human lung cancer A549 transplantation tumor in nude mice T/C (%) 3132 33 34 35 36 37 38 Sorafenib human lung 38.7 35.3 37.5 22.3 23.6 37.938.3 36.5 41.8 cancer A549 transplantation tumor Compound A′ Compound B′Compound C′ human lung 43.2 40.4 36.6 cancer A549 transplantation tumor

10. Tumor inhibition rate of tested compounds on human liver cancer cellbel-7402 transplantation tumor model in nude mice

[Test animals] female BALB/cA nude mice, 35-40 days old, with weight of18-22g. There were 12 mice in the negative conrol group and 6 mice inthe treatment group.

[Test method] Take eugenic tumor tissues and cut into about 1.5 mm³, andthen incoculate subcutaneously at the right armpit of nude mice underthe sterile conditions. The diameter of the transplantable tumor in nudemice was determined with a vernier caliper, and the animals were dividedinto groups after the tumors were grown to 100-300 mm³.Using the methodof measuring the tumor diameter, dynamically observe the antitumoreffects of tested materials.

The diameter of the tumor was determined three times every week and themouse weight was weighed at the same time. The dosage of the compoundwas 60 mg/kg, 6 times every week for 3 weeks. Sorafenib was oraladministrated with dosage of 60 mg/kg, 6 times every week for 3 weeks.Equal amount of normal saline was administrated in the negative controlgroup. Observe for one week after administration.

[Detection Indicators and Calculation Methods]

(1) Tumor volume (TV) is calculated as:

-   -   TV=1/2×a×b²    -   wherein a and b respectively represents length and width.

(2) Relative tumor volume (RTV) is calculated as:

-   -   RTV=TV_(t)/TV_(0°)

wherein TV₀ is the tumor volume when administrated according todifferent cages (d₀) and TV_(t) is the tumor volume measured each time.

(3) Relative tumor reproduction rate T/C (%) is calculated as follows:

${T\text{/}{C(\%)}} = {\frac{T_{RTV}}{C_{RTV}} \times 100}$

T_(RTV): RTV in the treatment group; C_(RTV): RTV in the negativecontrol group. The test results used relative tumor reproduction rateT/C (%) as evaluating indicator of anti-tumor activity.

[Screening results] There was no mortality for the animals in the groupof compounds and Sorafenib in the experiments with less toxicity.

TABLE 35 Relative tumor reproduction rate of compounds and Sorafenib onhuman liver cancer cell bel-7402 transplantation tumor model in nudemice T/C (%) Relative tumor reproduction rate of compounds 1-10 andSorafenib on human liver cancer cell bel-7402 transplantation tumormodel in nude mice T/C (%) 1 2 3 4 5 6 7 8 9 10 human liver 33.7 35.430.1 15.3 15.7 19.5 34.2 30.6 36.8 28.1 cancer cell bel-7402transplantation tumor 11 12 13 14 15 16 17 18 19 20 human liver 23.731.6 29.8 36.1 33.6 14.9 12.5 12.7 16.3 17.2 cancer cell bel-7402transplantation tumor 21 22 23 24 25 26 27 28 29 30 human liver 16.415.3 17.7 16.8 29.1 36.1 29.2 17.5 16.2 16.1 cancer cell bel-7402transplantation tumor 31 32 33 34 35 36 37 38 Sorafenib human liver 33.431.4 32.7 31.5 34.5 37.9 36.3 31.5 35.9 cancer cell bel-7402transplantation tumor Compound A′ Compound B′ Compound C′ human liver37.2 30.4 32.6 cancer cell bel-7402 transplantation tumor

11. Tumor inhibition rate of medicines on renal carcinoma cell lineGCR-1 transplanted tumor model in nude mice

[Test animals] female BALB/cA nude mice, 35-40 days old, with weight of18-22g. There were 12 mice in the negative conrol group and 6 mice inthe treatment group.

[Test method] Take eugenic tumor tissues and cut into about 1.5 mm³, andthen incoculate subcutaneously at the right armpit of nude mice underthe sterile conditions. The diameter of the transplantable tumor in nudemice was determined with a vernier caliper, and the animals were dividedinto groups after the tumors were grown to 100-300 mm³.Using the methodof measuring the tumor diameter, dynamically observe the antitumoreffects of tested materials. The diameter of the tumor was determinedthree times every week and the mouse weight was weighed at the sametime. The dosage of the medicine was 60 mg/kg, 6 times every week for 3weeks. Sorafenib was oral administrated with dosage of 60 mg/kg, 6 timesevery week for 3 weeks. Equal amount of normal saline was administratedin the negative control group. Observe for one week afteradministration.

[Detection Indicators and Calculation Methods]

(1) Tumor volume (TV) is calculated as:

-   -   TV=1/2×a×b²    -   wherein a and b respectively represents length and width.

(2) Relative tumor volume (RTV) is calculated as:

-   -   RTV=TV_(t)/TV_(0°)    -   wherein TV₀ is the tumor volume when administrated according to        different cages (d₀) and TV_(t) is the tumor volume measured        each time.

(3) Relative tumor reproduction rate T/C (%) is calculated as follows:

${T\text{/}{C(\%)}} = {\frac{T_{RTV}}{C_{RTV}} \times 100}$

-   -   T_(RTV): RTV in the treatment group; C_(RTV): RTV in the        negative control group. The test results used relative tumor        reproduction rate T/C (%) as evaluating indicator of anti-tumor        activity.

[Screening results] There was no mortality for the animals in the groupof compounds and Sorafenib in the experiments with less toxicity.

TABLE 36 Relative tumor reproduction rate of compounds and Sorafenib onhuman renal carcinoma GCR-1 cell transplanted tumor model in nude miceT/C (%) Relative tumor reproduction rate of compounds 11-20 andSorafenib on human renal carcinoma GCR-1 cell transplanted tumor modelin nude mice T/C (%) 1 2 3 4 5 6 7 8 9 10 human renal 30.5 23.2 31.3 9.611.2 12.5 21.2 29.1 28.3 27.1 carcinoma GCR-1 cell transplanted tumor 1112 13 14 15 16 17 18 19 20 human renal 32.1 20.5 28.3 31.1 22.5 9.9 11.112.3 13.2 13.1 carcinoma GCR-1 cell transplanted tumor Relative tumorreproduction rate of compounds 21-30 and Sorafenib on human renalcarcinoma GCR-1 cell transplanted tumor model in nude mice T/C (%) 21 2223 24 25 26 27 28 29 30 human renal 7.8 8.1 9.1 10.8 31.2 24.2 26.2 11.412.8 10.2 carcinoma GCR-1 cell transplanted tumor 31 32 33 34 35 36 3738 Sorafenib human renal 31.4 32.5 33.4 36.1 32.3 25.9 32.3 20.5 33.9carcinoma GCR-1 cell transplanted tumor Compound A′ Compound B′ CompoundC′ human renal 34.2 30.4 35.6 34.2 30.4 carcinoma GCR-1 celltransplanted tumor

According to the experimental results, the compound added with specificsubstituents in A ring have stronger anti-tumor activity than thecompouns with no substituent or only amino formyl in A ring, especiallythe 4#-6# 16#-18# 19#-24# 28#-30# compounds have stonger anti-tumoractivity which are stonger than the positive conrol Sorafenib, whichhave particularly evident effects on the tumor cell metastasis and tumorangiogenesis that are significantly stronger than Sorafenib. The test onnormal human umbilical vein endothelial cells CCK8 found that thesecompounds have less toxicity to normal human cells like endothelialcells, which are relatively safe and reliable, but these compounds canachieve the antitumor activity through inhibiting the tumorangiogenesis. The in vivo transplantation experiments in nude miceshowed that 4#-6# 16#-18# 19#-24# 28#-30# compounds have inhibitioneffects on human liver cancer and renal caner and their effects arebetter than Sorafenib, but these compounds have very significant effectson lung cancer and the effects obviously exceed the positive controlmedicine Sorafenib, which is an unexpected result.

The above results indicate that the compounds added with specificsubstituents in A-ring have more advantages than previously foundcompounds with no substituent or only amino formyl in A ring, and thesenew compounds have broader prospects in the treatment of cancer.

1. A compound or pharmaceutically acceptable salts thereof, wherein thecompound is represented by the following formula [1] or [2]

wherein, A is monosubstituted or polysubstituted quinoline,isoquinoline, quinazoline, pyrrole or pyrimidine, and the substituent ishalogen, C₁₋₅alkyl, C₁₋₅haloalkyl, C₁₋₅alkoxy, C₁₋₅haloalkoxy,C₁₋₅alkylamino, C₁₋₅haloalkylamino, amino or nitryl; R₁ is C₁₋₅alkyl; R₂is one or more selected from hydrogen, halogen, alkyl, alkoxy, haloalkylor haloalkoxy; and R₃ is one or more selected from hydrogen, halogen,alkyl, alkoxy, haloalkyl or haloalkoxy.
 2. The compound orpharmaceutically acceptable salts thereof according to claim 1, whereinA is monosubstituted or polysubstituted quinazoline, pyrrole orpyrimidine, and the substituent is halogen, C₁₋₅alkyl, C₁₋₅haloalkyl,C₁₋₅alkoxy, C₁₋₅haloalkoxy, C₁₋₅alkylamino, C₁₋₅haloalkylamino, amino ornitryl.
 3. The compound or pharmaceutically acceptable salts thereofaccording to claim 2, wherein A is monosubstituted or polysubstitutedquinazoline, and the substituent is halogen, C₁₋₅alkyl, C₁₋₅haloalkyl,C₁₋₅alkoxy, C₁₋₅haloalkoxy, amino or nitryl.
 4. The compound orpharmaceutically acceptable salts thereof according to claim 1, whereinR₁ is methyl or ethyl, propyl, isopropyl.
 5. The compound orpharmaceutically acceptable salts thereof according to claim 1, whereinR₂ is one or more selected from hydrogen, halogen, C₁₋₅alkyl, C₁₋₅alkoxy or C₁₋₅haloalkyl.
 6. The compound or pharmaceutically acceptablesalts thereof according to claim 5, wherein R₂ is one or more selectedfrom H, Cl, Br, F, methoxy, ethoxy, propoxy, methyl, ethyl, propyl,isopropyl, butyl, t butyl or trifluoromethyl.
 7. The compound orpharmaceutically acceptable salts thereof according to claim 1, whereinR₃ is one or more selected from hydrogen, halogen, C₁₋₅alkyl,C₁₋₅alkoxy, or C₁₋₅haloalkyl.
 8. The compound or pharmaceuticallyacceptable salts thereof according to claim 7, wherein R₃ is one or moreselected from H, Cl, Br, F, methoxy, ethoxy, propoxy, methyl, ethyl,propyl, isopropyl, butyl, t-butyl or trifluoromethyl.
 9. The compound orpharmaceutically acceptable salts thereof according to claim 1, whereinthe salts are selected from: a) basic salts of inorganic acids andorganic acids, the described acid is selected from hydrochloric acid,hydrobromic acid, sulfuric acid, phosphoric acid, mesylate,trifluoromethanesulfonic acid, benzene sulfonic acid,paratoluenesulfonic acid, 1-naphthalene sulfonic acid, 2-naphthalenesulfonic acid, acetic acid, trifluoroacetic acid, malic acid, tartaricacid, citric acid, lactic acid, oxalic acid, succinic acid, fumaricacid, maleic acid, benzoic acid, salicylic acid, phenylacetic acid oralmonds acid; b) acid salts of organic and inorganic base, the describedcation is selected from alkali metal cation, alkaline earth metalcation, ammonium cation, aliphatic-substituted ammonium cation oraromatic-substituted ammonium cation.
 10. The compound orpharmaceutically acceptable salts thereof according to any one of claims1 to 9, wherein the compound is selected from: the followingN′-(4-(2-(N-methylcarbamoyl)-4-(6-methoxyquinolinyl)oxy)phenyl)urea:N-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(6-methoxyquinolinyl)oxy)phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(6-methoxyquinolinyl)oxy)phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(6-methoxyquinolinyl)oxy)phenyl)urea;the following N′-(4-(2-(N-methylcarbamoyl)-4-(7-fluoroquinazolinyl)oxy)phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(7-fluoroquinazolinyl)oxy)phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(7-fluoroquinazolinyl)oxy)phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(7-fluoroquinazolinyl)oxy)phenyl)urea; the followingN′-(4-(2-(N-methylcarbamoyl)-5-(4-methoxypyrimidinyl)oxy)) phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-5-(4-methoxypyrimidinyl)oxy))phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-5-(4-methoxypyrimidinyl)oxy))phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-5-(4-methoxypyrimidinyl)oxy))phenyl)urea; the followingN′-(4-(2-(N-methylcarbamoyl)-4-(7-amino-isoquinolyl)oxy)phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(7-amino-isoquinolyl)oxy)phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(7-amino-isoquinolyl)oxy)phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(7-amino-isoquinolyl)oxy)phenyl)urea;the followingN′-(4-(2-(N-methylcarbamoyl)-4-(5-methyl-pyrryl)oxy))phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(5-methyl-pyrryl)oxy))phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(5-methyl-pyrryl)oxy))phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(2-(N-methylcarbamoyl)-4-(5-methyl-pyrryl)oxy))phenyl)urea;the following N′-(4-(4-(6-methoxyquinolinyl)oxy)phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy)phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy)phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyquinolinyl)oxy)phenyl)urea;the following N′-(4-(4-(7-fluoroquinazolinyl)oxy)phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-fluoroquinazolinyl)oxy)phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-fluoroquinazolinyl)oxy)phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-fluoroquinazolinyl)oxy)phenyl)urea;the following N′-(4-(5-(4-methoxypyrimidinyl)oxy))phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(5-(4-methoxypyrimidinyl)oxy))phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(5-(4-methoxypyrimidinyl)oxy))phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(5-(4-methoxypyrimidinyl)oxy))phenyl)urea;the following N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(7-amino-isoquinolyl)oxy)phenyl)urea;the following N′-(4-(4-(2-methyl-pyrryl)oxy))phenyl)ureaN-(4-chloro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(2-methyl-pyrryl)oxy))phenyl)urea;N-(4-fluoro-3-(trifluoromethyl)phenyl)-N′-(4-(4-(2-methyl-pyrryl)oxy))phenyl)urea;N-(4-bromo-3-(trifluoromethyl)phenyl)-N′-(4-(4-(2-methyl-pyrryl)oxy))phenyl)urea;the following substituted quinoline derivativesN-(4-chloro-3-ethylphenyl)-N′-(4-(4-(6-methoxyl-7-fluoro-quinolinyl)oxy)phenyl)urea;N-(4-ethyl-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methoxyl-7-fluoro-quinolinyl)oxy)phenyl)urea;N-(4-chloro-3-methoxylphenyl)-N′-(2-methyl-4-(2-(N-ethylcarbamoyl)-4-(7-nitryl-quinolinyl)oxy)phenyl)urea;N-(4-chloro-3-methoxylphenyl)-N′-(2-methoxyl-4-(2-(N-ethylcarbamoyl)-4-(7-nitryl-quinolinyl)oxy)phenyl)urea;the following substituted quinazoline derivativesN-(4-chloro-3-ethylphenyl)-N′-(4-(4-(6-methyl-7-fluoro-quinazolinyl)oxy)phenyl)urea;N-(4-ethyl-3-(trifluoromethyl)phenyl)-N′-(4-(4-(6-methyl-7-fluoro-quinazolinyl)oxy)phenyl)urea;N-(4-chloro-3-methoxylphenyl)-N′-(2-fluoro-4-(2-(N-propylcarbamoyl)-4-(7-trifluoromethylquinazolinyl)oxy)phenyl)urea;N-(4-chloro-3-methoxylphenyl)-N′-(2-trifluoromethyl-4-(2-(N-propylcarbamoyl)-4-(7-trifluoromethylquinazolinyl)oxy)phenyl)urea.11. A method of treating tumor or leukemia in subject, the methodcomprising administering the subject an effective dosage amount of thecompound or pharmaceutically acceptable salts thereof according to anyone of claims 1 to 9.